Parathyroid hormone (PTH) enhances the osteogenic response of bone to mechanical loads, in vivo. We postulate that PTH interacts with the signaling mechanisms of mechanotransduction to lower the mechanical threshold and prime the osteogenic cells of bone to respond to lesser magnitudes of mechanical stimulation to promote bone formation. We have shown that PTH can alter both mechanosensitive (MSCC) and L-type voltage sensitive calcium channel (LVSCC) kinetics to increase the Ca2+ signal in osteoblasts, resulting in increased osteogenic activity. Further, we have shown that reorganization of the actin cytoskeleton leads to increased activity of MSCC and that PTH-induced activation of protein kinase A (PKA) enhances activity of both the L-VSCC and the MSCC. We have also shown that inhibition of the L-VSCC significantly reduces the bone formation rate in both rats and mice subjected to mechanical loading and completely blocks the PTH enhanced bone formation response to in vivo loading. Our overall hypothesis is that PTH reduces the mechanical threshold in osteoblasts by altering MSCC and L-VSCC kinetics to increase the entry of Ca2+ in response to a mechanical signal. In this proposal, we will continue to examine the mechanisms of PTH control of the mechanotransduction signaling pathways and how this regulation alters cell function. The aims of this proposal are: 1) examine the role of the cytoskeleton in the regulation of MSCC and L-VSCC kinetics and intracellular Ca2+ release in osteoblasts in response to shear and ascertain how PTH modulates this regulation, 2) determine the effects of phosphorylation on the regulation of the MSCC and L-VSCC kinetics fluid shear and PTH stimulation in osteoblasts and 3) ascertain how PTH enhanced Ca2+'entry through the MSCC and L-VSCC alters release of autocrine/paracrine factors and increase gene expression in osteoblasts. Completion of these aims should provide insight into the regulation of bone by the mechanical environment and how modulation of the signaling pathways can affect bone formation.